V-ATPase在TMEM25调节溶酶体酸化及癫痫发作中的作用及机制研究

癫痫发作导致神经元溶酶体功能紊乱。前期研究发现，跨膜蛋白25（TMEM25）是调节溶酶体酸化的重要蛋白，过表达TMEM25可抑制兴奋性突触传递和减少癫痫发作，但其作用机制尚不清楚。溶酶体是自噬和内吞成分的降解中枢，液泡膜-ATP酶（V-ATPase）介导的质子在溶酶体酸化中起重要作用。通过预实验发现，TMEM25与V-ATPase存在相互作用，可影响V-ATPase表达水平，抑制V-ATPase可阻断TMEM25对神经元活性的作用。据此我们提出假说：V-ATPase介导的质子转运在TMEM25调节溶酶体酸化及癫痫发作中起重要作用。本课题拟在分子、细胞和动物等多层面，利用分子生物学、全细胞膜片钳、动物行为学等技术，明确TMEM25对于V-ATPase介导的质子转运的调节作用，从而揭示TMEM25调节癫痫发作及溶酶体酸化的机制，为进一步探索TMEM25的功能及抗癫痫治疗的新靶点提供理论依据。

Epileptic seizures lead to neuronal lysosome dysfunction. In our previous study, we have found that transmembrane protein 25 (TMEM25) is an important protein regulating lysosomal acidification. Overexpression of TMEM25 can inhibit excitatory synaptic transmission and reduce epileptic seizures. But its mechanism is unclear. Lysosomes are the degradation centers of autophagy and endocytosis components, and V-ATPase is a pH-sensitive proton transporter that plays an important role in lysosomal acidification. It was found that TMEM25 interacts with V-ATPase, could regulate the expression level of V-ATPase and inhibition of V-ATPase could block the effect of TMEM25 on neuronal activity. Based on this, we hypothesized that V-ATPase mediated proton transport plays an important role in TMEM25's regulation of lysosomal acidification and epileptic seizures. This study intends to clarify the effect of TMEM25 on V-ATPase mediated proton transport and lysosomal acidification, and its mechanism on the levels of molecule, cell, animal and tissues from epileptic patients, by using molecular biology, whole-cell patch clamp and animal experiments techniques. By these studies, we hope to reveal the mechanism of TMEM25 regulating epileptic seizures and to provide a new theoretical basis for exploring new targets for antiepileptic.